Coopting the Lap system ofPseudomonas fluorescensto reversibly customize bacterial cell surfaces

Abstract

Initialattachment to a surface is a key and highly regulated step in biofilm formation. In this study, we present a platform for reversibly functionalizing bacterial cell surfaces, with an emphasis on designing biofilms. We engineered the Lap system of Pseudomonas fluorescens Pf0-1, which is normally used to regulate initial cell surface attachment, to display various protein cargo at the bacterial cell surface and control extracellular release of the cargo in response to changing levels of the second messenger cdi-GMP. To accomplish this goal, we fused the protein cargo between the N-terminal retention module and C-terminal secretion signal of LapA, and controlled surface localization of the cargo with natural signals known to stimulate or deplete c-di-GMP levels in P. fluorescens Pf0-1. We show this system can tolerate large cargo in excess of 500 amino acids, direct P. fluorescens Pf0-1 to surfaces it does not typically colonize, and program this microbe to sequester the toxic medal cadmium.

Engineering biofilm matrix for efficient biofilm-mediated contaminant removal

“…With an aim to facilitate the molecular cloning procedures, this study explores the feasibility of truncating BpfA down to its essential genetic component that maintains the S. oneidensis biofilm formation. Smith et al has demonstrated how a repeats-in-toxin (RTX) adhesin, LapA (520 kDa), can be truncated to act as a protein cargo to carry cadmium binding domain by retaining its N-terminal retention module and C-terminal secretion module [255]. Since…”